The overall Aim of this Project will be to conduct clinical vaccine trials in humans that advance our understanding of vaccine induced mucosal immunity. The studies are linked closely to the ongoing macaques and murine investigations described in Projects 1 and 2. During the first year of the Program will be responsible for developing the protocols, submitting the appropriate regulatory paperwork, and recruiting the subjects needed to develop and validate mucosal assays. Samples from HIV-positive patients and seronegative controls will be collected to develop HIV antigen-specific mucosal assays. In the second year we will validate assays using samples from mucosal and central compartments by conducting a trial of a live-attenuated influenza virus vaccine delivered by intranasal and intrarectal routes. The goal of this vaccine trial will be in part to generate T cell immune responses that can be measured both in the periphery and mucosal compartments and to compare and correlate these responses. In months 36-48, we will carry out a dose ranging trial of Sindbis virus ogp140deltaV2.SF162, administered by the specific mucosal route determined in the first 2 years of the Program in the murine and primate projects (Projects 1 and 2). For the sake of trial design in this application, we assume that intranasal administration will be equal or superior to immunizing via either the intrarectal routes in males or the intravaginal route in females, in terms of inducing protection from disease (in the macaque model) after either intrarectal or intravaginal challenge. In the final year of the Project we will carry out a prime boast systemic/mucosal vaccine trial based on the cumulative results obtained in the first four years of the overall Program. The Project 4 clinical program will closely interface with Projects 1-3, the Vaccine Technology Core (VTC), and the Administrative Core to develop these protocols. This planned vaccine trail in humans will be conducted concomitantly with a mucosal in rhesus macaques, which will use the same vaccine delivered by the same routes at the same doses.